Journal: Neural Regeneration Research

Article Title: Mitogen activated protein kinase signaling pathways participate in the active principle region of Buyang Huanwu decoction-induced differentiation of bone marrow mesenchymal stem cells ☆

doi:

Figure Lengend Snippet: CD11b, CD45 and CD90 expression in rat bone marrow mesenchymal stem cells detected by flow cytometry. (A) Blank control cells. (B-D) FITC labeled CD90, CD11 and CD45 positive cells respectively. FITC: Fluorescein isothiocyanate.

Article Snippet: Cells were then incubated with fluorescein isothiocyanate-labeled anti-CD11b, -CD45 and -CD90 antibodies (1:1 000; 0.5 mL; SERATEC, Gottingen, Germany).

Techniques: Expressing, Flow Cytometry, Control, Labeling

Journal: Glia

Article Title: RvE1 treatment prevents memory loss and neuroinflammation in the Ts65Dn mouse model of Down syndrome

doi: 10.1002/glia.23779

Figure Lengend Snippet: Antibody details

Article Snippet: The membrane was incubated with secondary antibodies conjugated with either IRDye 680RD or 800CW (Li-COR Biosciences, Lincoln, NE) for 30 min and then washed with TBS, 3 × 5 min. All probe signals were measured with a Li-COR Odyssey scanner and quantified with Image Studio Lite (Li-COR Biosciences). table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Antibody Manufacturer Method Dilution Rb anti-leukotriene B4 receptor 1 (BLT1) Cayman Chemical, Ann Arbor, MI Immunoblot 1:1,000 Ms anti-chemokine-like receptor 1 (ChemR23) Santa Cruz Biotechnology, Dallas, TX Immunoblot 1:200 Rb anti-Peroxisome proliferator-activated receptor gamma (PPAR-γ) R&D Systems, Minneapolis, MN Immunoblot 1:1,000 Rb anti-N-formyl peptide receptor 2 (LxA4R) Santa Cruz Biotechnology, Dallas TX Immunoblot 1:500 Rb anti-phospho (Thr202/Tyr204) extracellular-signal-regulated kinase (Erk1/2) Cell Signaling, Danvers, MA Immunoblot 1:1,000 Ms anti-total extracellular-signal-regulated kinase (Erk1/2) Cell Signaling, Danvers, MA Immunoblot 1:1,000 Rb anti-15-lipoxygenase-2 (15LOX2) Cayman Chemical, Ann Arbor, MI Immunoblot 1:200 Rb anti-calbindin-D28 Bio-Rad, Hercules, CA Immunoblot 1:200 Rb anti-ionized calcium binding adaptor molecule 1 (Iba1) MilliporeSigma, Burlington, MA Immunohistochemistry 1:200 Rb anti-protein tyrosine phosphatase, receptor type, C (CD45) Wako-Chem, Richmond, VA Immunohistochemistry 1:200 Open in a separate window Antibody details

Techniques: Western Blot, Binding Assay, Immunohistochemistry

Journal: Glia

Article Title: RvE1 treatment prevents memory loss and neuroinflammation in the Ts65Dn mouse model of Down syndrome

doi: 10.1002/glia.23779

Figure Lengend Snippet: Microglial inflammatory response to RvE1 treatment. (a) Typical CD45-staining seen in Veh-treated NS mice showed multiple resting microglia with small cell somas and long, thin processes, which were not affected by RvE1 treatment (b). (c) Veh-treated Ts65Dn mice displayed elevated CD45-staining of microglial cells displaying an activated morphology compared to the NS mice (a). (d) RvE1 treatment reversed the microglial CD45 phenotype of Ts65Dn mice to staining patterns observed in NS cohorts. Scale bar equals 500 μm. (e) Two-way ANOVA effects by karyotype and RvE1 treatment for CD45 densitometry confirmed that RvE1 treatment significantly reduced CD45 staining density in Ts65Dn mice. (f) RvE1 treatment also reversed elevated 15-LOX-2 levels in the hippocampus of Ts65Dn mice. Two-way ANOVA effects by karyotype and RvE1 treatment of 15-LOX-2 densitometry confirmed that RvE1 treatment significantly reduced 15-LOX-2 levels in Ts65Dn mice. (g) Calbindin-D28 was significantly reduced in Ts65Dn mice and not affected by RvE1 treatment. Two-way ANOVA effects by karyotype only on averaged calbindin-D28 densitometry confirmed that RvE1 treatment had no effects on calbindin-D28 levels in Ts65Dn mice. Tukey’s post hoc p values are shown for group comparisons and error bars represent mean ± SEM. (h) CD45 density correlated significantly with spontaneous locomotion (positive correlation), total WRAM errors (positive correlation), and with NORT performance (negative correlation). Dotted lines represent 95% confidence interval. CD45, protein tyrosine phosphatase, receptor type, C; 15-LOX-2, 15-Lipoxygenase-2; NS, normosomic; NORT-DI, novel object recognition discrimination index; RvE1, resolvin E1; TS, Ts65Dn; Veh, vehicle; WRAM, water radial arm maze task

Article Snippet: The membrane was incubated with secondary antibodies conjugated with either IRDye 680RD or 800CW (Li-COR Biosciences, Lincoln, NE) for 30 min and then washed with TBS, 3 × 5 min. All probe signals were measured with a Li-COR Odyssey scanner and quantified with Image Studio Lite (Li-COR Biosciences). table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Antibody Manufacturer Method Dilution Rb anti-leukotriene B4 receptor 1 (BLT1) Cayman Chemical, Ann Arbor, MI Immunoblot 1:1,000 Ms anti-chemokine-like receptor 1 (ChemR23) Santa Cruz Biotechnology, Dallas, TX Immunoblot 1:200 Rb anti-Peroxisome proliferator-activated receptor gamma (PPAR-γ) R&D Systems, Minneapolis, MN Immunoblot 1:1,000 Rb anti-N-formyl peptide receptor 2 (LxA4R) Santa Cruz Biotechnology, Dallas TX Immunoblot 1:500 Rb anti-phospho (Thr202/Tyr204) extracellular-signal-regulated kinase (Erk1/2) Cell Signaling, Danvers, MA Immunoblot 1:1,000 Ms anti-total extracellular-signal-regulated kinase (Erk1/2) Cell Signaling, Danvers, MA Immunoblot 1:1,000 Rb anti-15-lipoxygenase-2 (15LOX2) Cayman Chemical, Ann Arbor, MI Immunoblot 1:200 Rb anti-calbindin-D28 Bio-Rad, Hercules, CA Immunoblot 1:200 Rb anti-ionized calcium binding adaptor molecule 1 (Iba1) MilliporeSigma, Burlington, MA Immunohistochemistry 1:200 Rb anti-protein tyrosine phosphatase, receptor type, C (CD45) Wako-Chem, Richmond, VA Immunohistochemistry 1:200 Open in a separate window Antibody details

Techniques: Staining

Journal: Nature Communications

Article Title: iCLOTS: open-source, artificial intelligence-enabled software for analyses of blood cells in microfluidic and microscopy-based assays

doi: 10.1038/s41467-023-40522-4

Figure Lengend Snippet: Designed for use with fluorescence microscopy images such that multiple components of a cell suspension can be simultaneously monitored, users adjust threshold values for any image color channel(s) where immunofluorescence signal is present. a Line graphs representing occlusion and accumulation over time are automatically generated. Here, CD41+ platelets and CD45+ white blood cells from sickle cell disease patient whole blood accumulate on an ibidi chamber device coated with collagen at a faster rate than healthy control whole blood ( n = 3 replicates). Data taken at ×20 magnification, scale bar represents 50 μm. b The application is designed to automatically generate a map of all signals present, e.g., the dimensions of a microfluidic device, shown here with a microvasculature-on-a-chip device designed to investigate the effect of crizanlizumab on sickle cell disease whole blood samples ( n = 1 experiment). Data taken at ×20, scale bar represents 200 μm. Percent occlusion ( c ) and accumulation rate ( d ) changes over the course of an experiment, showing microvascular occlusion instability. e A microchannel-specific application is available for spatial analysis of one or many straight microchannel portions of a microfluidic device. Data taken at ×20, scale bar represents 50 μm. f Spatial quantification of microvascular occlusion is automatically performed by calculating an occlusion percentage for each pixel point along the length of each microchannel. ML algorithms enable further analysis in microchannels by treating each x-coordinate and corresponding occlusion measurement from each channel as a data point. At the initial time course timepoint, t = 7 min, CD45+ white blood cells in SCD whole blood ( g ) and CD45+ white blood cells in SCD whole blood treated with drug crizanlizumab ( h ) occlude endothelialized microchannels to variable degrees at each point along the 32 analyzed microchannels. i CD45+ white blood cells in SCD whole blood predominantly occlude distal ends of microchannels at early timepoints, while CD45+ WBCs in SCD whole blood treated with crizanlizumab occlude proximal entry points of microchannels at early timepoints. Source data are provided as a Source data file.

Article Snippet: In order to facilitate perfusion, syringes were loaded with whole blood treated with 40 µg/ml corn trypsin inhibitor (CTI; Haematologic Technologies), Anti-CD45 Mouse Monoclonal Antibody (VWR) at a ratio of 1:100, Integrin alpha 2b/CD41 Antibody (VWR) at a ratio of 1:200, and 6 mM CaCl 2 .

Techniques: Fluorescence, Microscopy, Immunofluorescence, Generated

Journal: Nature Communications

Article Title: iCLOTS: open-source, artificial intelligence-enabled software for analyses of blood cells in microfluidic and microscopy-based assays

doi: 10.1038/s41467-023-40522-4

Figure Lengend Snippet: a Analysis windows are designed to be intuitively followed from left (inputs) to right (outputs), with the image processing steps as applied displayed in the center. Here, the microchannel analysis application from iCLOTS’ suite of multiscale microfluidic accumulation tools is shown with whole blood perfused through an in vitro microvasculature-on-a-chip microfluidic model. b The user uploads the desired number of microscopy images, time course microscopy series, or videomicroscopy files as inputs. These files are then automatically displayed on the screen. c Depending on the application and file type, users are guided through a series of windows facilitating the analyses of their data, such as choosing a region of interest (ROI, shown) or indicating immunofluorescence staining color channels present in a file. iCLOTS applications designed for fluorescence microscopy can accommodate up to three stains in separate channels: here, red indicates CD41+ platelets, green indicates CD45+ white blood cells, and blue indicates the endothelial cell layer. Data in this example is taken at ×100 magnification, scale bars represent 50 μm (left) and 10 μm (right). d Parameters, numerical factors that define how image processing algorithms should be applied, are typically simple, e.g., minimum and maximum cell area, or fluorescence signal threshold, as shown here. All parameters are adjusted interactively from a default value to best match the researcher’s specific dataset. In iCLOTS, pixel intensity values are understood to be arbitrary units. Effects of changing parameters are shown in real time to assist in gauging the appropriateness of selected values. e A button initiates the finalized analysis with algorithms customized by the selected parameters. Upon completion, graphical results appropriate for the application are automatically displayed, such as line graphs representing quantitative accumulation and occlusion values at each time point for each channel of interest, as seen in this example. f Users may export any of the outputs generated by iCLOTS, including tabular data as an Excel file, graphical results as .png images, or the initial imaging dataset as transformed by the image processing algorithms and/or labeled with indices.

Article Snippet: In order to facilitate perfusion, syringes were loaded with whole blood treated with 40 µg/ml corn trypsin inhibitor (CTI; Haematologic Technologies), Anti-CD45 Mouse Monoclonal Antibody (VWR) at a ratio of 1:100, Integrin alpha 2b/CD41 Antibody (VWR) at a ratio of 1:200, and 6 mM CaCl 2 .

Techniques: In Vitro, Microscopy, Immunofluorescence, Staining, Fluorescence, Generated, Imaging, Transformation Assay, Labeling